As recently reviewed, SERT-deficient Slc6a4 +/- and -/- mice have gene-proportionate increases in extracellular fluid serotonin (5-HT) concentrations, i.e., 3- or 6-fold excesses respectively over +/+ mice (Murphy et al., 2008;Murphy &Lesch 2008;Fox et al., 2007, 2009, 2010;Brigman et al., 2010). At the same time, the Slc6a4 -/- mice have a marked deficit of intracellular, releasable 5-HT (Fox et al., 2008). These mice also have significantly increased 5-HT synthesis and turnover across multiple brain regions investigated in Slc6a4 -/- mice. Likewise, Slc6a4 +/- mice manifest decreased 5-HT clearance and elevated extracellular fluid 5-HT but nonetheless have unchanged tissue 5-HT concentrations in the brain and periphery and unchanged brain 5-HT synthesis and turnover. Thus, a single copy of Slc6a4 is adequate to maintain overall 5-HT tissue homeostasis, although other phenotypes such as cocaine preference and adverse reactions to pro-serotonergic agents are clearly different in Slc6a4 partially-deficient +/- mice. Continuing advances have been made in our studies of the serotonin-related toxic reactions, including the serotonin syndrome. Most commonly, this toxicity occurs as a side effect in humans treated with certain antidepressant and anti-anxiety drugs. Importantly, its milder forms contribute to reduced therapeutic efficacy or a requirement to interrupt treatment in some individuals treated with SRIs. Our earlier studies exploring this behavioral and temperature-related syndrome in SERT-deficient mice had discovered that genetic vulnerability to a markedly exaggerated serotonin syndrome was present when these mice were exposed to the metabolic precursor of serotonin, 5-HTP, or to other serotonergic drugs such as the monoamine oxidase inhibiting (MAOI) antidepressant, tranylcypromine. Tranylcypromine is a clinically available antidepressant drug, and 5-HTP is readily available on more than 100 internet sites and sold over-the-counter as a dietary supplement for the treatment of depression, insomnia and related disorders. Spontaneous mild serotonin-syndrome behaviors were also observed in untreated Slc6a4 -/- mice, a reflection of the demonstrated markedly increased serotonin availability in these mice (Fox et al., 2007;Fox et al., 2008). This past year, we have expanded our serotonin syndrome studies and demonstrated an exaggerated response to atypical opiods including meperidine and tramadol. Given this transgenic mice data and our other human SLC6A4 polymorphism data, we have formed an international collaborative effort - "Genes Involved in SEroTonin Toxicity" or "GISETTO" - in which we are examining functional variants in the multiple serotonin (SLC6A4, MAOA) and drug metabolizing gene groups (e.g., CYP2D6) genes in individuals developing serotonin toxicity and other serotonin-related toxicities, both retrospectively and prospectively. Understanding these CNS genetic mechanisms will improve serotonergic drug safety and efficacy by identifying patients whose response to drugs may be compromised, and thereby be at risk of adverse and potentially life threatening drug reactions. SERT provides an intriguing example of likely mouse-human congruence in genetic vulnerability to serotonin toxicity features. Functional variants exist in additional genes that can also be postulated to confer vulnerability to serotonin neurotoxicity, for example, monoamine oxidase and serotonin receptor genes. Exaggerated changes in temperature and other pharmacological responses following serotonin-enhancing drugs were also found in SERT-deficient mice. Different serotonin receptor mechanisms were discovered to be involved in the altered responses by studies with different selective serotonin antagonists and additional agonists, including the 5-HT2A receptor agonist, TCB-2 and the anti-pain medication, tramadol. In SERT +/+ and +/- mice, the 5-HT1A autoreceptor was primarily involved. However, in SERT -/- mice, the 5-HT7 receptor was the primary mediator of temperature changes (Fox et al., 2008). As roles for 5-HT7 receptors in anxiety and depression were recently established and contributions of 5-HT1A and other 5-HT receptors to therapeutic responses to serotonergic drugs have been well-established, the current findings have implications for understanding the high anxiety-like and depressive-like phenotype of SERT-deficient mice that are relevant to the treatment of human anxiety and affective disorders. These findings in this mouse genetic model raise the possibility that humans with lower-expressing SLC6A4 SS genotypes or other SERT variants that lead to 50-80% decreases in SERT binding sites or transport function may be at higher risk to develop serotonin syndrome neurotoxicity. This is based on highly congruent data from imaging, neuroendocrine and other studies that have compared SERT-deficient mice, and SERT-deficient non-human primates to humans with SERT gene variants. Of special note, it is likely that relatively mild serotonin syndrome occurrence may contribute to early discontinuation of SRIs and other side effects during SRI treatment that are strongly associated with the lower-expressing SLC6A4 SS genotypes and S alleles as well as other newly discovered lower-expressing variants in this gene, as described in our other report, MH000336-30 LCS. SERT-deficient mice show reduced head twitch responses to serotonergic agents such as the 5-HT2A receptor agonist and hallucinogen, DOI. This response is considered a pharmacologic proxy for hallucinogenic-like behaviors, as most of the drugs that induce this response in rodents are hallucinogens in humans, and these responses can be blocked by anti-psychotic drugs. In exploring mechanisms underlying this response, we found that reducing excess brain serotonin in SERT-deficient mice by using a serotonin synthesis inhibitor restored the deficient head twitch state in SERT -/- mice to levels of SERT control +/+ mice (Basselin et al., 2009). These findings provide a more comprehensive understanding of mechanisms in this genetic mouse model that eventually may provide insights into human disorders with genetic contributions that include hallucinogenic phenomena such as schizophrenia and bipolar disorder as well as the consequences of certain types of substance abuse disorders--all of high impact, major public health concern. SERT-deficient mice were also compared to mice with gene-based deficiencies in other neurotransmitters, those for dopamine and norepinephrine, as well as to mice with other genetic deficiencies that interact with SERT, such as the integrin alphaIIbeta3 gene, as well as other types of environmental stimuli. Differences in brain, platelet and gut responses to multiple agents were observed. These findings serve as further illustrations of earlier data accumulated, as referenced below, by our Lab that supports the use of these mice as vulnerability models for humans with SERT gene variants with regard to gene-environment and gene-gene interactions that contribute to human diseases and their pharmacologic treatment. In additional studies of SERT-deficient mice, these mice were examined for possible autism-related traits such as social interaction deficiencies. Some studies have identified associations of SLC6A4 polymorphisms as well as plasma serotonin levels with the occurrence of autism. In a three-chambered choice task, male SERT-deficient mice displayed significantly reduced social interaction behaviors with other mice compared to SERT +/+ mice (Moy et al., 2009). Thus, together with increased anxiety-like behaviors plus increased startle responses and increased behavioral and physiological responses to various types of stress, these mice show a series of features highly relevant to symptoms in several neuropsychiatric disorders of important public concern.